Meiosis is a specialized cell cycle essential in all species for the generation of functional gametes. The female gamete, or oocyte, becomes arrested early in development typically during meiosis I, and must be reactivated at a later time point in order to ensure fertilization. This reactivation is termed oocyte meiotic maturation and is required in all female species. We are using C. elegans to investigate the mechanisms behind oocyte meiotic maturation. Our lab has previously shown that an inhibitory kinase of the cell cycle, WEE-1.3, is required for proper timing of oocyte meiotic maturation. Upon depletion of WEE-1.3 via RNA interference oocytes mature precociously and although the oocytes encounter sperm, they are not fertilized and the worm becomes sterile. The mechanism by which wee-1.3 RNAi results in precocious oocyte maturation, potential genetic interactors with wee-1.3 and the subcellular localization of wild type WEE-1.3 remain unknown. To begin to address these questions we generated transgenic worms containing a translational fusion of WEE-1.3 to GFP. We found that WEE-1.3 exhibits perinuclear localization throughout the germline and distinct punctae in many of the cells comprising the multicellular embryo. We are currently performing colocalization studies in attempt to describe the WEE-1.3 expression pattern in more detail. In order to determine genes that interact with wee-1.3, we performed an RNAi suppressor screen utilizing 1874 embryonic lethal clones from the OpenBiosystems RNAi library. From this screen we identified 150 genes that when depleted in conjunction with depletion of WEE-1.3 result in a restoration of fertility. We have begun to confirm the identified suppressors and to characterize the mechanisms through which they suppress the sterility of wee-1.3 RNAi, which includes analyzing the germlines of suppressed worms to determine the state of oocyte maturation. We will also report on whether any suppressors are affecting the localization of CDK-1, a substrate of WEE-1.3. The strength of all these approaches is its potentiality for identifying novel components of both the cell cycle and oocyte meiotic maturation.

Affiliation:
- LBG, NIDDK, National Institutes of Health, Bethesda, MD